Small G-proteins (RAC and RHO) are known to be involved in regulation of superoxide (O2•–) production and the assembly of actin fibres. These processes are known to be crucial for accessibility and inaccessibility of barley cells to the biotrophic fungus Blumeria graminis f.sp. hordei (Bgh). Using a candidate RT-PCR approach six Rac-related cDNA-clones were isolated from barley. The transient knock-down of RacB led to a remarkably lower penetration efficiency of Bgh into susceptible barley lines (Mlo/Ror1). Surprisingly the inhibition of RacB expression had no effect in the double mutant line A89 (mlo5/ror1). This led us to the assumption that the RacB-dsRNA effect is dependent on functional ROR1 (such as mlo5-mediated resistance). Vice versa, overexpression of constitutive active RACB-V15 in the susceptible line Pallas resulted in hypersusceptibility to Bgh. Thus, we conclude that RACB is a signal transduction protein functional in the accessibility of epidermal barley cells to Bgh.